The behavioral pharmacological profile of a drug in a pertinent species is necessary for evaluating quantitatively not only how the drug functions as a reinforcer but also how use or abuse of the drug affects other aspects of the subject's behavior. Ongoing studies on the direct behavioral effects of drugs include a number of different paradigms, including two-lever, choice, drug-discrimination behavior, operant behavior maintained by food or by postponement of avoidance of electric shock, and in-vivo microdialysis measurements of neurotransmitter levels in discrete brain areas. In one series of studies we are using laboratory animals to characterize interactions of caffeine with the effects of other psychomotor stimulants under conditions of acute and chronic caffeine exposure. Caffeine and nicotine are the main psychoactive ingredients of coffee and tobacco, with a high frequency of concurrent use in humans. In previous experiments we demonstrated that there are qualitative differences in behavioral response to psychomotor stimulant drugs which result from chronic exposure to caffeine and depend on the level of caffeine exposure. In a recent study we further examined the interaction of caffeine and nicotine under a drug-discrimination paradigm in rats trained to discriminate nicotine from saline under a fixed ratio schedule of food presentation. One group of rats was chronically exposed to caffeine (1.0 mg/ml) dissolved in their drinking bottles whereas the other group was exposed to tap water. Effects of intraperitoneal injections of caffeine on nicotine-lever selection were subsequently examined. In separate groups of rats exposed to the same caffeine-drinking or water-drinking regimen, effects of caffeine pretreatment on nicotine blood levels were evaluated. Although caffeine (1.0 - 30.0 mg/kg) did not generalize to nicotine when administered alone, it markedly potentiated discriminative-stimulus effects of the threshold dose of nicotine in both water- and caffeine-drinking animals. Nicotine blood levels were, however, not affected by acute or chronic caffeine exposure. Thus, caffeine enhances the discriminative effects of nicotine by a pharmacodynamic rather than pharmacokinetic interaction. We have also found that caffeine given either acutely or chronically potentiates behavioral effects of the direct and indirect dopaminergic agonists, amphetamine and cocaine. This is thought to reflect caffeine's ability to bind to postsynaptic adenosine receptors, preventing antagonistic effects of adenosine on dopaminergic neurotransmission. In a recent series of in vivo microdialysis experiments, we found that chronic caffeine exposure enhances the increases in extracellular concentrations of dopamine in the shell of the nucleus accumbens produced by injection of nicotine or metahamphetamine. The nucleus accumbens is the ventral part of the striatum and is an area involved in the motor and motivational effects of psychostimulant drugs. A strong antagonistic interaction between subtypes of adenosine and dopamine receptors has been found in the striatum. Adenosine A2A receptors interact with dopamine D2 receptors and adenosine A1 receptors interact with dopamine D1 receptors. Also glutamate is thought to interact with dopamine and adenosine systems and modulate some of the effects of drugs acting on these two systems. We have now demonstrated, for the first time, that behaviorally relevant doses of caffeine significantly increase extracellular levels of both dopamine and glutamate in the shell of the nucleus accumbens. Peak increases of about 100% above baseline with dopamine and about 50% above baseline with glutamate occurred. These findings show that presynaptic mechanisms are also involved in the effects of caffeine. Furthermore these presynaptic effects are mostly mediated by A1 receptors. We are currently investigating the effects of selective A1 and A2 receptor antagonists on dopamine and glutamate neurotransmission in the nucleus accumbens of rats and comparing them to those of caffeine to further elucidate the involvement of presynaptic mechanisms in the enhancement of the behavioral effects of psychomotor stimulants by acute and chronic caffeine exposure. To analyze postsynaptic mechanisms of adenosine receptor signaling in the striatum, which are mostly mediated by adenosine A2A receptors, we have initiated a series of experiments using immunohistochemical techniques, to evaluate the A2A receptor mediated modulation of the striatal expression of the immediate-early gene c-fos. We have so far demonstrated that A2A receptor signaling depends on co-stimulation of a subtype of glutamate metabotropic receptor (mGluR5) and on co-stimulation of presynaptic A1 receptors. As a consequence of these studies, in collaboration with Rafael Franco (Barcelona) and Amina Woods (NIDA), we have demonstrated the existence of heteromeric complexes between A2A and mGluR5 recceptors. Together with our recent demonstration of A2A/D2 receptor heteromeric complexes, caffeine should be contemplated as a drug that targets multimeric receptor complexes, which allows us to begin to understand the complex and contradictory pharmacological actions of this drug.